Looking for ALPS: The value of a combined assessment of biochemical markers.

BACKGROUND: Autoimmune lymphoproliferative syndrome (ALPS) is a rare primary immune disorder caused by defect of the extrinsic apoptotic pathway. The current diagnostic criteria combine clinical features and typical biomarkers but have not been the object of clear international consensus. METHODS: We conducted a retrospective study on pediatric patients who were investigated for autoimmune cytopenia and/or lymphoproliferation at the CHU Sainte-Justine Hospital over 10 years. Patients were screened using the combination of TCRαß+ CD4- CD8- "double negative" (DN) T cells and soluble plasmatic FAS ligand (sFASL). RESULTS: Among the 398 tested patients, the median sFASL and DN T cells were 200 ng/mL and 1.8% of TCRαß+ T cells, respectively. sFASL was highly correlated with vitamin B12 levels. We identified five patients diagnosed with ALPS for whose sFASL and vitamin B12 levels were the more discriminating biomarkers. While ALPS diagnostic criteria had high sensibility, their predictive value remained low. CONCLUSION: sFASL level can efficiently discriminate patients with ALPS when using the appropriate thresholds. Our study highlights the need for an international consensus to redefine the place and threshold of biological biomarkers for ALPS diagnosis.

PI3Kδ coordinates transcriptional, chromatin, and metabolic changes to promote effector CD8+ T cells at the expense of central memory.

Patients with activated phosphatidylinositol 3-kinase delta (PI3Kδ) syndrome (APDS) present with sinopulmonary infections, lymphadenopathy, and cytomegalvirus (CMV) and/or Epstein-Barr virus (EBV) viremia, yet why patients fail to clear certain chronic viral infections remains incompletely understood. Using patient samples and a mouse model (Pik3cdE1020K/+ mice), we demonstrate that, upon activation, Pik3cdE1020K/+ CD8+ T cells exhibit exaggerated features of effector populations both in vitro and after viral infection that are associated with increased Fas-mediated apoptosis due to sustained FoxO1 phosphorylation and Fasl derepression, enhanced mTORC1 and c-Myc signatures, metabolic perturbations, and an altered chromatin landscape. Conversely, Pik3cdE1020K/+ CD8+ cells fail to sustain expression of proteins critical for central memory, including TCF1. Strikingly, activated Pik3cdE1020K/+ CD8+ cells exhibit altered transcriptional and epigenetic circuits characterized by pronounced interleukin-2 (IL-2)/STAT5 signatures and heightened IL-2 responses that prevent differentiation to memory-like cells in IL-15. Our data position PI3Kδ as integrating multiple signaling nodes that promote CD8+ T cell effector differentiation, providing insight into phenotypes of patients with APDS.

Quantitative assessment of the relationship between Fas/FasL genes polymorphisms and head and neck cancer risk.

BACKGROUND: Molecular epidemiological studies have demonstrated a closer association between Fas/FasL polymorphisms and head and neck cancer (HNC) risk, and the results of these published studies were inconsistent. We therefore performed this meta-analysis to explore the associations between Fas/FasL polymorphisms and HNC risk. METHODS: Four online databases (PubMed, Embase, CNKI, and Wanfang) were searched. Odds ratios (ORs) with 95% confidence interval (95% CIs) were calculated to assess the association between Fas -670A>G, Fas -1377G>A, and FasL -844C>T polymorphisms and HNC risk. In addition, heterogeneity, accumulative/sensitivity analysis, and publication bias were conducted to check the statistical power. RESULTS: Overall, 9 related publications (20 independent case-control studies) involving 3179 patients and 4217 controls were identified. Significant association of protective effects was observed between FasL -844C>T polymorphism and HNC risk in codominant and dominant model models (CT vs CC: OR = 0.89, 95% CI = 0.79-1.00, P = .05, I = 38.3%, CT+TT vs CC: OR = 0.88, 95% CI = 0.79-0.98, P = .02, I = 35.8%). Furthermore, the similar protective effects were observed the subgroup analysis of in Asian population and population-based controls group. CONCLUSION: Our meta-analysis indicated that FasL -844C>T polymorphism plays a protective role against HNC development, but the Fas -670A>G and Fas -1377G>A polymorphisms maybe not associated with HNC risk.

Quantitative assessment of the association between Fas/FasL gene polymorphism and susceptibility to esophageal carcinoma in a north Chinese population.

The case-control study aims to investigate the association of Fas and FasL genetic polymorphisms (Fas-670A/G (rs1800682), Fas-1377G/A (rs2234767) and FasL-844T/C (rs763110)) with esophageal carcinoma susceptibility in a north Chinese population. A total of 204 patients with esophageal carcinoma and 248 healthy controls were enrolled from Henan, China and genotyped by the polymerase chain reaction and restriction fragment length polymorphism method. There were no significant differences in distributions of their genotypes frequencies between patients and controls in Fas-670A/G, Fas-1377G/A and FasL-844T/C polymorphisms (P > 0.05). Stratified analysis showed that no significant association was found between esophageal carcinoma and gene polymorphisms of Fas-670 A/G, Fas-1377G/A, and FasL-844T/C (P > 0.05). Genetic polymorphisms in the death pathway genes Fas and FasL were not associated with risk of developing esophageal carcinoma in a north Chinese population.

Apoptotic and anti-apoptotic seromarkers for assessment of disease severity of non-alcoholic steatohepatitis.

BACKGROUND AND STUDY AIMS: This study aimed to find out non-invasive markers for the assessment of severity of non-alcoholic steatohepatitis (NASH) in an attempt to decrease the need for liver biopsy. It also aimed to evaluate the key role of apoptosis in the pathogenesis of the disease and the suggested role of anti-apoptotic factors in therapeutic modalities and disease prognosis. PATIENTS AND METHODS: The serum levels of soluble Fas (s. Fas), s. Fas ligand, cytokeratin 18 (CK-18) fragment and Bcl-2 were measured in 80 patients and 15 non-hepatic subjects as control. The patients were divided based on histological examination of liver biopsy into three groups. Group I included 40 patients with NASH, group II had 40 patients with non-alcoholic fatty liver disease (NAFLD) non-NASH and group III had 15 non-hepatic subjects as control. Apoptosis of hepatocytes was assessed by morphological examination using a light microscope and expressed as number per square millimetre. RESULTS: There was a significant increase in the serum levels of s. Fas, s. Fas ligand and CK-18 fragments in the NASH group. The anti-apoptotic protein Bcl-2 showed significantly low levels in NASH patients. Apoptosis of hepatocytes was significantly higher in the NASH group. The degree of apoptosis was inversely correlated with the level of Bcl-2. A significant correlation between both s. Fas and CK-18 fragment with liver histology with regard to lobular inflammation and ballooning was found. CONCLUSIONS: Increased serum levels of s. Fas and CK-18 fragment in the NASH group and its correlation with the severity of disease suggested the key role of apoptosis in NASH pathogenesis which can be used for the assessment of the severity of NASH. A high level of anti-apoptotic Bcl-2 in NAFLD suggests its protective role in disease progress.

Quantitative assessment of the association between three polymorphisms in FAS and FASL gene and breast cancer risk.

FAS and FAS ligand (FASL) play crucial roles in apoptotic signaling, and deregulation of this pathway may facilitate carcinogenesis. Studies on the association between the FAS/FASL polymorphisms (FAS-1377G/A rs2234767, FAS-670A/G rs1800682, and FASL-844C/T rs763110) and breast cancer risk have reported inconsistent results. Therefore, to characterize the relationship between those polymorphisms and breast cancer risk, we undertook a meta-analysis of those studies. Several electronic databases were searched for articles on the FAS/FASL polymorphisms and breast cancer risk. The genotype data were extracted; pooled odds ratios (OR) with 95% confidence intervals (CIs) were used to estimate the strength of the association. Five studies were eligible for our meta-analysis. Overall, we observed significant associations of the FAS-1377G/A polymorphism with breast cancer susceptibility (AG vs. GG: OR = 1.15, 95% CI 1.02-1.30; AA vs. GG: OR = 1.39, 95% CI 1.12-1.72; AG/AA vs. GG: OR = 1.18, 95% CI, 1.16-1.32; A vs. G: OR = 1.16, 95% CI 1.06-1.26), but we did not observe significant association of the Fas-670A/G and FasL-844C/T polymorphisms with breast cancer risk. In the subgroup analysis, we observed that the FAS-1377G/A and FASL-844C/T polymorphisms were associated with breast cancer risk in Chinese but not Whites; we still did not observed association of the FAS-670A/G polymorphism with breast cancer risk. Our meta-analysis revealed that FAS-1377G>A polymorphism was associated with an increased risk of breast cancer. FASL-844C>T polymorphism might be associated with a reduced breast cancer risk in Chinese. However, FAS-670A/G had no any effect on breast carcinogenesis.

Methyl helicterate protects against CCl4-induced liver injury in rats by inhibiting oxidative stress, NF-κB activation, Fas/FasL pathway and cytochrome P4502E1 level.

This study was designed to investigate the protective effects of the methyl helicterate (MH) isolated from Helicteres angustifolia L. against CCl4-induced hepatotoxicities in rats. Liver injury was induced in rats by the administration of CCl4 twice a week for 8 weeks. Compared with the CCl4 group, MH significantly decreased the activities of ALT, AST and ALP in the serum and increased the activities of SOD, GSH-Px and GSH-Rd in the liver. Moreover, the content of hepatic MDA was reduced. Histological findings also confirmed the anti-hepatotoxic characterisation. In addition, MH significantly inhibited the proinflammatory mediators, such as PGE2, iNOS, COX-2, IL-6, TNF-α and myeloperoxidase (MPO). Further investigation showed that the inhibitory effect of MH on the proinflammatory cytokines was associated with the downregulation of NF-κB. Besides, MH also markedly decreased the levels of Fas/FasL protein expression and the activities of caspase-3/8, as well as the activity of cytochrome P4502E1 (CYP2E1). In brief, the protective effect of MH against CCl4-induced hepatic injury may rely on its ability to reduce oxidative stress, suppress inflammatory responses, protect against Fas/FasL-mediated apoptosis and block CYP2El-mediated CCl4 bioactivation.

[Role of proteins in Fas-mediated apoptosis in tumor cells and lymphocytes co-cultured in vitro].

The investigation deals with the role of Fas, FasL, RIP, caspase 3, and PARP taking part in Fas-mediated apoptosis, and contributing to in vitro interaction of hepatoma MH-22a and histiocytic sarcoma J-774 in mice with syngenic splenocytes. Protein expression was identified by means of indirect immunofluorescence. There were two patterns of interaction of tumor cells and splenocytes: apoptosis occurred either in 80% or in an insignificant number of tumor cells. In the latter case, high Fas expression was identified before and when it dropped after the experiment. FasL expression in tumor cells often peaked before the experiment and then it decreased after contact with lymphocytes. That mechanism was reversed in splenocytes: contact with tumor cells boosted expression. RIP, caspase 3 and PARP expression was very low and failed to show until the experiments on both patterns of cells were undertaken. After the experiments, it either remained latent or soared up. In the latter case, simultaneous expression of all proteins took place both in tumor cells and lymphocytes. A second battery of experiments demonstrated maximum rates of apoptosis both of tumor cells and splenocytes. However, the situation was different: Fas expression intensified in both patterns of cells after their interaction which was followed by post-experimental drop in RIP, caspase 3, and PARP expression in tumor cells; hence, the importance of perforin/granzyme-mediated apoptosis which occurred at the early stages of tumor growth in the midst of interaction with immune system cells. That pattern of apoptosis was highly cytotoxic. It is suggested that Fas-mediated apoptosis or any other receptor-sensitive pathway might take place during tumor progression, i.e. at a stage when tumor is most susceptible to change.

The role of apoptosis in respiratory diseases.

PURPOSE: The purpose of this article is to define apoptosis and describe how this cellular pathway is relevant to the pathogenesis of different respiratory diseases. This will assist clinical nurse specialists in understanding how new drugs and therapies inhibit and stimulate apoptotic pathways. BACKGROUND: Clinical nurse specialists need to expand their knowledge concerning the role of apoptosis so that they can better expand their spheres of influence. The 4 stages of apoptosis are discussed, as well as the various apoptotic pathways involved with asthma, emphysema, and acute respiratory distress syndrome that promote and inhibit apoptosis in patients. CONCLUSION: It is crucial for clinical nurse specialists to know what apoptosis is and how it relates to different pathophysiologic states. The challenge facing clinical nurse specialists is how to be kept informed and current concerning molecular and cellular mechanisms that are important in the practice setting. Strategies needed to maintain expertise include acquiring new knowledge, developing new skills, and changing attitudes about molecular biology. Apoptosis must become a significant part of any health professionals' continuing educational program because it has been recognized as the pathway to most any disease. Clinical nurse specialists who understand apoptosis and its pathways can use this knowledge to aid in the prevention and treatment of respiratory diseases.

Integrated mechanistic and data-driven modelling for multivariate analysis of signalling pathways.

Mathematical models of highly interconnected and multivariate signalling networks provide useful tools to understand these complex systems. However, effective approaches to extracting multivariate regulation information from these models are still lacking. In this study, we propose a data-driven modelling framework to analyse large-scale multivariate datasets generated from mathematical models. We used an ordinary differential equation based model for the Fas apoptotic pathway as an example. The first step in our approach was to cluster simulation outputs generated from models with varied protein initial concentrations. Subsequently, decision tree analysis was applied, in which we used protein concentrations to predict the simulation outcomes. Our results suggest that no single subset of proteins can determine the pathway behaviour. Instead, different subsets of proteins with different concentrations ranges can be important. We also used the resulting decision tree to identify the minimal number of perturbations needed to change pathway behaviours. In conclusion, our framework provides a novel approach to understand the multivariate dependencies among molecules in complex networks, and can potentially be used to identify combinatorial targets for therapeutic interventions.

[The RT-PCR-identified Fas and FasL expression and apoptosis in mouse hepatoma MH-22a and histiocytic sarcoma J-774 clonal lines coupled with in vitro cultivation with syngenic splenocytes].

Malignant growth is associated with various patterns of interaction between tumor cells and those of the body immune system, interaction between Fas-receptor (Fas) and Fas-ligand (FasL) expression being one of them. These mechanisms were simulated in vitro using the main cell populations from murine hepatoma MH-22a, histiocytic sarcoma J-774 and their clonal lines obtained from cocultivation of tumor cells and syngenic splenocytes. Fas and FasL expression was identified by the RT-PCR method while apoptosis--by electrophoresis of low molecular DNA fractions and clonogenic survival.

Pancreatic beta and alpha cells are both decreased in patients with fulminant type 1 diabetes: a morphometrical assessment.

AIMS/HYPOTHESIS: We have previously reported that fulminant type 1 diabetes is characterised by an absence of diabetes-related antibodies and a remarkably abrupt onset. However, little is known about the mechanism of beta cell destruction in this diabetes subtype, and to obtain insights into the aetiology of the disease, we investigated residual endocrine cells and the expression of Fas and Fas ligand in fulminant type 1 diabetes. METHODS: Residual beta and alpha cells were morphologically assessed in pancreatic tissue obtained by biopsy from five patients with recent-onset fulminant type 1 diabetes and five patients with recent-onset typical autoimmune type 1 diabetes. In addition, the expression of Fas and Fas ligand was evaluated by immunohistochemistry. RESULTS: In fulminant type 1 diabetes, beta and alpha cell areas were decreased significantly, compared with autoimmune type 1 diabetes and control subjects. In contrast, the alpha cell area was not decreased significantly in autoimmune type 1 diabetes, compared with that in control subjects. No Fas expression in islets and Fas ligand expression in CD3(+) cells in the exocrine pancreas were found in the fulminant type 1 diabetic patients who underwent this evaluation. CONCLUSIONS/INTERPRETATION: Our study showed that beta and alpha cells are damaged in fulminant type 1 diabetes. In addition to the lack of Fas and Fas ligand expression, the results suggest that the mechanism of beta cell destruction in fulminant type 1 diabetes is different from that in autoimmune type 1 diabetes.

Assessment of cytotoxic lymphocyte gene expression in the peripheral blood of human islet allograft recipients: elevation precedes clinical evidence of rejection.

Studies in nonhuman primates have demonstrated that elevation of the cytotoxic lymphocyte (CL) genes granzyme B, perforin, and Fas ligand in peripheral blood precedes islet allograft rejection. The purpose of this study was to determine whether this approach has utility for prediction of human islet allograft loss. We studied 13 patients who had long-term type 1 diabetes and were treated with steroid-free immunosuppression and given sequential islet cell infusions. All recipients became insulin independent, and eight of them experienced deterioration in glycemic control, followed by reinitiation of insulin therapy. Frequent peripheral blood samples were collected to monitor CL gene mRNA levels with real-time PCR. For the eight back-to-insulin patients, there was a clear elevation of CL gene mRNA levels 25-203 days before the onset of frequent hyperglycemia. Granzyme B was the most reliable indicator of ongoing graft loss. Additional correlations with infection were noted; however, evidence of sensitization in antidonor mixed lymphocyte reaction was observed in seven of eight patients who experienced partial graft loss, whereas this was not seen when upregulated CL gene expression was associated with infection. The results suggest that, when taken into consideration with other clinical parameters, elevated CL gene levels may enable prediction of islet allograft loss.

Apoptosis in proliferative vitreoretinopathy.

PURPOSE: To study the involvement of apoptosis using different apoptosis markers in PVR pathogenesis. METHODS: The presence of mRNA coding for Fas, Fas ligand (FasL), and TNF-related apoptosis inducing ligand (TRAIL) was investigated in vitreous samples from 46 consecutive patients-25 with PVR, 11 with retinal detachment (RD) not complicated by PVR, and 10 with macular hole (MH)-using RT-PCR. From previously examined vitreous samples, 21 PVR, 9 RD, and 10 MH were examined for their levels of TGF-beta2 protein with sandwich ELISA kits. Five epiretinal membranes excised from five patients with PVR were also examined for apoptotic cell death using the terminal deoxytransferase (TdT) mediated dUTP-biotin nick end labeling (TUNEL) technique. RESULTS: FAS mRNA was detected in 72% of patients with PVR, 55% of patients with RD and 20% of patients with MH. TRAIL mRNA was detected in 67% of patients with PVR, 89% of patients with RD, and 20% of patients with MH. FasL mRNA was detected in 20% of patients with PVR, 9% of patients with RD, and 10% of patients with MH. The median levels of Fas and TRAIL mRNA were significantly higher (P < 0.05) in patients with PVR than in those with MH hole but between patients with PVR and those with RD the difference was not significant (P > 0.05). A significant difference was detected between RD and MH for TRAIL mRNA levels (P = 0.008). For FasL, no significant difference between groups was found. TGF-beta2 was detected in all investigated vitreous samples. A significant difference was found between the PVR and MH groups (P = 0.001) and between the RD and MH groups (P = 0.004), but not between the PVR and RD groups (P < 0.05). The level of TGF-beta2 was significantly correlated to the level of TRAIL mRNA (r = 0.86), but no correlation was found between TGF-beta2 and Fas mRNA levels (r = 0.21). Four of five examined PVR epiretinal membranes showed positive staining for apoptotic cells using the TUNEL technique. CONCLUSIONS: Apoptosis is one of the mechanisms that is involved in PVR pathogenesis. Different apoptosis markers suggest different pathways occur in PVR, including Fas/FasL, TRAIL, and TGF-beta2 mediated processes.

Induction of salivary gland epithelial cell injury in Sjogren's syndrome: in vitro assessment of T cell-derived cytokines and Fas protein expression.

Sjogren's syndrome (SS) is an exocrinopathy characterized by T cell infiltrates, salivary gland epithelial cell (SGEC) apoptosis and high Fas and FasL expression. To address the participation of T cell-derived cytokines and of Fas apoptotic pathway in SS glandular lesions, we utilized non-neoplastic SGEC lines established from SS patients and controls. Possibly attesting to their intrinsic activation, cell lines derived from SS patients displayed significantly higher constitutive Fas and FasL than controls. Surface co-expression of Fas and FasL was not associated with spontaneous fratricide apoptosis. SGEC were resistant to anti-Fas-mediated apoptosis (possibly owing to the constitutive expression of anti-apoptotic proteins cFLIP and Bcl-2), but became sensitive after protein or RNA synthesis inhibition. IFN-gamma and TNF-alpha were able to upregulate surface Fas and FasL, whereas IL-1beta downregulated surface FasL. IFN-gamma (but not several other cytokines) reduced the survival of SGEC in a dose- and time-dependent manner and induced Fas/FasL-mediated apoptosis, directly and via anoikia. Dexamethasone inhibited the upregulation of Fas and FasL by IFN-gamma and the induction of SGEC apoptosis and detachment by anti-Fas mAb or IFN-gamma. Our findings indicate the injurious role of IFN-gamma for the salivary epithelia of SS patients through the induction of Fas-mediated apoptosis and anoikia.

Soluble fas levels correlate with multiple organ dysfunction severity, survival and nitrate levels, but not with cellular apoptotic markers in critically ill patients.

Apoptosis is a mode of programmed cell death (PCD). Transduction of apoptotic signals results in cellular suicide. Organ specific apoptosis has been proposed as a factor in multiple organ dysfunction syndrome (MODS). Fas is a widely occurring apoptotic signal receptor molecule expressed by almost any type of cell, which is also released in a soluble circulating form (circulating fas, sfas). In this exploratory study, we investigated the association of sfas with severity, survival, known mediators of multiple organ dysfunction, and cellular apoptotic markers on peripheral blood mononuclear cells (PBMC) in a group of 35 patients with MODS and in 35 matched controls. Critically ill patients with MODS had significantly elevated sfas levels compared to controls over time (P < .001). Increased serum concentration of circulating fas was associated with increased severity of multiple organ dysfunction. Non-survivors exhibited significantly higher sfas levels compared to survivors (P < .01) and increasing sfas was inversely associated with the likelihood of survival (P < .05). Circulating fas levels correlated highly with serum nitrate concentration, but not with fas and fasL expression on PBMC of critically ill patients. TNF-alpha and IL-6, although they appear to be mediators of both apoptosis and MODS, had no association with sfas. These results are suggestive of the need for further investigation on the role of apoptotic signaling in the development of MODS. They also suggest a potential prognostic value of sfas for SIRS/MODS clinical outcomes.